A procedure which has proven successful in identifying dominant acting oncogenes associated with various human malignancies has been DNA transfection (1-3). By employing DNA Ca.sup.2+ -mediated DNA transfection techniques and various molecular cloning strategies, the genetic elements presumably responsible for malignant conversion of specific human and rodent cells have been identified and cloned (4-21). The primary method used to identify transforming oncogenes has been by Ca.sup.2+ -mediated DNA transfer of sheared HMW-tumor DNA into NIH-3T3 cells followed by the isolation and characterization of DNA from morphologically transformed foci. A limitation of this assay has been that the majority of HMW-DNA from primary and established human tumor isolates, approximately 80%, fail to induce morphological transformation of NIH-3T3 cells.
By using a modification of this procedure, the Ca.sup.2+ -mediated transfer of HMW-DNA plus the neomycin resistance gene into NIH-3T3 cells, selection for neomycin resistant colonies in G418-supplemented medium, injection of pooled neomycin resistant cells into nude mice and the generation and subsequent isolation of tumor cells, it has been possible to identify transforming DNA which either fails to form morphologically transformed foci or is weakly focus-producing when transfected into NIH-3T3 cells (1,12,13).
More recent studies indicate that a specific clone of Fisher rat embryo cells, CREF (14), develops a tumorigenic phenotype in nude mice after being cotransfected with high molecular weight (HMW-prostatic carcinoma DNA or HMW-colon carcinoma DNA in combination with pSV2-Neo plasmid DNA followed by selection for G418 resistant colonies (2). In contrast, both types of HMW-DNAs did not induce foci of morphologically altered CREF cells when assayed using a monolayer culture system (2). In addition, when the same HMW tumor-DNA samples and transfection protocol used with CREF cells were applied to NIH-3T3 cells, no tumors were induced by transfected cells in nude mice (2).
The recent development of hybridoma technology has resulted in the development of specific monoclonal antibodies to various human tumor associated antigens. This approach has greatly facilitated the study of the potential role of tumor associated antigens in expression of the neoplastic state (38). However, the production of monoclonal antibodies with good tumor specificity, especially toward various stages in the development of a malignant neoplasm, remains a laborious and uncertain undertaking, especially when freshly isolated heterogeneous tumor cells are used as immunogens. An innovative recent approach to the generation of monoclonal antibodies with specificity to transforming gene products which may be mediators of the tumor state, has been to utilize transfected heterologous species cells containing and expressing human transforming gene products as immunogens (39-42). Using this approach, monoclonal antibodies have been produced against the neu transforming gene transferred and expressed in NIH-3T3 cells. Similarly, Roth, et al. have used the transfection approach in generating monoclonal antibodies against NIH-3T3 transfectants produced following transfer of HMW-DNA from a human acute lymphocytic leukemia cell line (ALL) and to a c-Ha-ras transformed cell line (40, 41). In addition, Hollingsworth, et al. transfected NIH-3T3 cells with HMW-DNA from a human pancreatic adenocarcinoma cell line to generate monoclonal antibodies (42).